Genetic suppression of the lethality of rad53 null-mutation in Saccharomyces cerevisiae by overexpression of Cyc8 protein

초록

CYC8 (CRT8) was originally identified as a trans-acting crt (constitutive RNR3 transcription) mutant that express high levels of RNR3 in the absence of DNA damaging agents. It forms a complex with Tup1 and acts as a general transcriptional co-repressor. It also acts as part of a transcriptional co-activator complex that recruits the SWI/SNF and SAGA complexes to promoters. Rad53 is an essential checkpoint kinase that plays a central role in cell cycle arrest at all checkpoints. It also functions in transcriptional activation in response to DNA damage. RNR, the key enzyme in the maintenance and regulation of dNTP biosynthesis, is one of the target genes for transcriptional control by Rad53. Therefore, the hydroxyurea-sensitivity of rad53 mutant yeast cells is most likely to be caused by failure of regulation of dNTP levels. In this study, we screened multi-copy suppressor genes that suppress the hydroxyurea sensitivity of rad53 mutant, and identified CYC8 as a candidate gene. Overexpression of CYC8 by a multi-copy plasmid suppressed the lethality of rad53 null-mutation. The yeast cells transformed with this plasmid showed higher dNTP levels than were observed in the cells with control plasmid. These results suggest that elevation of the intracellular dNTP levels renders RAD53 dispensable for yeast cell viability.